Regulation of IGF1R by MicroRNA-15b Contributes to the Anticancer Effects of Calorie Restriction in a Murine C3-TAg Model of Triple-Negative Breast Cancer.

Calorie restriction (CR) inhibits triple-negative breast cancer (TNBC) progression in several preclinical models in association with decreased insulin-like growth factor 1 (IGF1) signaling. To investigate the impact of CR on microRNAs (miRs) that target the IGF1/IGF1R pathway, we used the spontaneous murine model of TNBC, C3(1)/SV40 T-antigen (C3-TAg). In C3-TAg mice, CR reduced body weight, IGF1 levels, and TNBC progression. We evaluated the tumoral expression of 10 miRs. CR increased the expression of miR-199a-3p, miR-199a-5p, miR-486, and miR-15b. However, only miR-15b expression correlated with tumorigenicity in the M28, M6, and M6C C3-TAg cell lines of TNBC progression. Overexpressing miR-15b reduced the proliferation of mouse (M6) and human (MDA-MB-231) cell lines. Serum restriction alone or in combination with low levels of recombinant IGF1 significantly upregulated miR-15b expression and reduced Igf1r in M6 cells. These effects were reversed by the pharmacological inhibition of IGFR with BMS754807. In silico analysis using miR web tools predicted that miR-15b targets genes associated with IGF1/mTOR pathways and the cell cycle. Our findings suggest that CR in association with reduced IGF1 levels could upregulate miR-15b to downregulate Igf1r and contribute to the anticancer effects of CR. Thus, miR-15b may be a therapeutic target for mimicking the beneficial effects of CR against TNBC.

Oxygen availability influences the incidence of testicular teratoma in Dnd1Ter/+ mice.

Testicular teratomas and teratocarcinomas are the most common testicular germ cell tumors in early childhood and young men, and they are frequently found unilaterally in the left testis. In 129/SvJ mice carrying a heterozygous copy of the potent modifier of tumor incidence Ter, a point mutation in the dead-end homolog one gene (Dnd1 Ter/+), ∼70% of the unilateral teratomas arise in the left testis. We previously showed that in mice, left/right differences in vascular architecture are associated with reduced hemoglobin saturation and increased levels of the hypoxia inducible factor-1 alpha (HIF-1α) in the left compared to the right testis. To test the hypothesis that systemic reduction of oxygen availability in Dnd1 Ter/+ mice would lead to an increased incidence of bilateral tumors, we placed pregnant females from 129/SvJ Dnd1 Ter/+ intercross matings in a hypobaric chamber for 12-h intervals. Our results show that in 129/SvJ Dnd1 Ter/+ male gonads, the incidence of bilateral teratoma increased from 3.3% to 64% when fetuses were exposed to acute low oxygen conditions for 12-h between E13.8 and E14.3. The increase in tumor incidence correlated with the maintenance of high expression of pluripotency genes Oct4, Sox2 and Nanog, elevated activity of the Nodal signaling pathway, and suppression of germ cell mitotic arrest. We propose that the combination of heterozygosity for the Ter mutation and hypoxia causes a delay in male germ cell differentiation that promotes teratoma initiation.

The role of SPAG1 in the assembly of axonemal dyneins in human airway epithelia.

Mutations in SPAG1, a dynein axonemal assembly factor (DNAAF) that facilitates the assembly of dynein arms in the cytoplasm before their transport into the cilium, result in primary ciliary dyskinesia (PCD), a genetically heterogenous disorder characterized by chronic oto-sino-pulmonary disease, infertility and laterality defects. To further elucidate the role of SPAG1 in dynein assembly, we examined its expression, interactions and ciliary defects in control and PCD human airway epithelia. Immunoprecipitations showed that SPAG1 interacts with multiple DNAAFs, dynein chains and canonical components of the R2TP complex. Protein levels of dynein heavy chains (DHCs) and interactions between DHCs and dynein intermediate chains (DICs) were reduced in SPAG1 mutants. We also identified a previously uncharacterized 60 kDa SPAG1 isoform, through examination of PCD subjects with an atypical ultrastructural defect for SPAG1 variants, that can partially compensate for the absence of full-length SPAG1 to assemble a reduced number of outer dynein arms. In summary, our data show that SPAG1 is necessary for axonemal dynein arm assembly by scaffolding R2TP-like complexes composed of several DNAAFs that facilitate the folding and/or binding of the DHCs to the DIC complex.

Expression of a Truncated Form of ODAD1 Associated with an Unusually Mild Primary Ciliary Dyskinesia Phenotype.

Primary ciliary dyskinesia (PCD) is a rare lung disease caused by mutations that impair the function of motile cilia, resulting in chronic upper and lower respiratory disease, reduced fertility, and a high prevalence of situs abnormalities. The disease is genetically and phenotypically heterogeneous, with causative mutations in > 50 genes identified, and clinical phenotypes ranging from mild to severe. Absence of ODAD1 (CCDC114), a component of the outer dynein arm docking complex, results in a failure to assemble outer dynein arms (ODAs), mostly immotile cilia, and a typical PCD phenotype. We identified a female (now 34 years old) with an unusually mild clinical phenotype who has a homozygous non-canonical splice mutation (c.1502+5G>A) in ODAD1. To investigate the mechanism for the unusual phenotype, we performed molecular and functional studies of cultured nasal epithelial cells. We demonstrate that this splice mutation results in the expression of a truncated protein that is attached to the axoneme, indicating that the mutant protein retains partial function. This allows for the assembly of some ODAs and a significant level of ciliary activity that may result in the atypically mild clinical phenotype. The results also suggest that partial restoration of ciliary function by therapeutic agents could lead to significant improvement of disease symptoms.

Mechanistic Targets and Nutritionally Relevant Intervention Strategies to Break Obesity-Breast Cancer Links.

The worldwide prevalence of overweight and obesity has tripled since 1975. In the United States, the percentage of adults who are obese exceeds 42.5%. Individuals with obesity often display multiple metabolic perturbations, such as insulin resistance and persistent inflammation, which can suppress the immune system. These alterations in homeostatic mechanisms underlie the clinical parameters of metabolic syndrome, an established risk factor for many cancers, including breast cancer. Within the growth-promoting, proinflammatory milieu of the obese state, crosstalk between adipocytes, immune cells and breast epithelial cells occurs via obesity-associated hormones, angiogenic factors, cytokines, and other mediators that can enhance breast cancer risk and/or progression. This review synthesizes evidence on the biological mechanisms underlying obesity-breast cancer links, with emphasis on emerging mechanism-based interventions in the context of nutrition, using modifiable elements of diet alone or paired with physical activity, to reduce the burden of obesity on breast cancer.

Induction of ciliary orientation by matrix patterning and characterization of mucociliary transport.

Impaired mucociliary clearance (MCC) is a key feature of many airway diseases, including asthma, bronchiectasis, chronic obstructive pulmonary disease, cystic fibrosis, and primary ciliary dyskinesia. To improve MCC and develop new treatments for these diseases requires a thorough understanding of how mucus concentration, mucus composition, and ciliary activity affect MCC, and how different therapeutics impact this process. Although differentiated cultures of human airway epithelial cells are useful for investigations of MCC, the extent of ciliary coordination in these cultures varies, and the mechanisms controlling ciliary orientation are not completely understood. By introducing a pattern of ridges and grooves into the underlying collagen substrate, we demonstrate for the first time, to our knowledge, that changes in the extracellular matrix can induce ciliary alignment. Remarkably, 90% of human airway epithelial cultures achieved continuous directional mucociliary transport (MCT) when grown on the patterned substrate. These cultures maintain transport for months, allowing carefully controlled investigations of MCC over a wide range of normal and pathological conditions. To characterize the system, we measured the transport of bovine submaxillary gland mucin (BSM) under several conditions. Transport of 5% BSM was significantly reduced compared with that of 2% BSM, and treatment of 5% BSM with the reducing agent tris(2-carboxyethyl)phosphine (TCEP) reduced viscosity and increased the rate of MCT by approximately twofold. Addition of a small amount of high-molecular-weight DNA increased mucus viscosity and reduced MCT by ∼75%, demonstrating that the composition of mucus, as well as the concentration, can have significant effects on MCT. Our results demonstrate that a simple patterning of the collagen substrate results in highly coordinated ciliated cultures that develop directional MCT, and can be used to investigate the mechanisms controlling the regulation of ciliary orientation. Furthermore, the results demonstrate that this method provides an improved system for studying the effects of mucus composition and therapeutic agents on MCC.

Mutation of CFAP57, a protein required for the asymmetric targeting of a subset of inner dynein arms in Chlamydomonas, causes primary ciliary dyskinesia.

Primary ciliary dyskinesia (PCD) is characterized by chronic airway disease, reduced fertility, and randomization of the left/right body axis. It is caused by defects of motile cilia and sperm flagella. We screened a cohort of affected individuals that lack an obvious axonemal defect for pathogenic variants using whole exome capture, next generation sequencing, and bioinformatic analysis assuming an autosomal recessive trait. We identified one subject with an apparently homozygous nonsense variant [(c.1762C>T), p.(Arg588*)] in the uncharacterized CFAP57 gene. Interestingly, the variant results in the skipping of exon 11 (58 amino acids), which may be due to disruption of an exonic splicing enhancer. In normal human nasal epithelial cells, CFAP57 localizes throughout the ciliary axoneme. Nasal cells from the PCD patient express a shorter, mutant version of CFAP57 and the protein is not incorporated into the axoneme. The missing 58 amino acids include portions of WD repeats that may be important for loading onto the intraflagellar transport (IFT) complexes for transport or docking onto the axoneme. A reduced beat frequency and an alteration in ciliary waveform was observed. Knockdown of CFAP57 in human tracheobronchial epithelial cells (hTECs) recapitulates these findings. Phylogenetic analysis showed that CFAP57 is highly conserved in organisms that assemble motile cilia. CFAP57 is allelic with the BOP2/IDA8/FAP57 gene identified previously in Chlamydomonas reinhardtii. Two independent, insertional fap57 Chlamydomonas mutant strains show reduced swimming velocity and altered waveforms. Tandem mass tag (TMT) mass spectroscopy shows that FAP57 is missing, and the "g" inner dyneins (DHC7 and DHC3) and the "d" inner dynein (DHC2) are reduced, but the FAP57 paralog FBB7 is increased. Together, our data identify a homozygous variant in CFAP57 that causes PCD that is likely due to a defect in the inner dynein arm assembly process.

Identification of genetic variants in CFAP221 as a cause of primary ciliary dyskinesia.

Primary ciliary dyskinesia (PCD) is a rare disorder that affects the biogenesis or function of motile cilia resulting in chronic airway disease. PCD is genetically and phenotypically heterogeneous, with causative mutations identified in over 40 genes; however, the genetic basis of many cases is unknown. Using whole-exome sequencing, we identified three affected siblings with clinical symptoms of PCD but normal ciliary structure, carrying compound heterozygous loss-of-function variants in CFAP221. Computational analysis suggests that these variants are the most damaging alleles shared by all three siblings. Nasal epithelial cells from one of the subjects demonstrated slightly reduced beat frequency (16.5 Hz vs 17.7 Hz, p = 0.16); however, waveform analysis revealed that the CFAP221 defective cilia beat in an aberrant circular pattern. These results show that genetic variants in CFAP221 cause PCD and that CFAP221 should be considered a candidate gene in cases where PCD is suspected but cilia structure and beat frequency appear normal.

Lack of GAS2L2 Causes PCD by Impairing Cilia Orientation and Mucociliary Clearance.

Primary ciliary dyskinesia (PCD) is a genetic disorder in which impaired ciliary function leads to chronic airway disease. Exome sequencing of a PCD subject identified an apparent homozygous frameshift variant, c.887_890delTAAG (p.Val296Glyfs∗13), in exon 5; this frameshift introduces a stop codon in amino acid 308 of the growth arrest-specific protein 2-like 2 (GAS2L2). Further genetic screening of unrelated PCD subjects identified a second proband with a compound heterozygous variant carrying the identical frameshift variant and a large deletion (c.867_∗343+1207del; p.?) starting in exon 5. Both individuals had clinical features of PCD but normal ciliary axoneme structure. In this research, using human nasal cells, mouse models, and X.laevis embryos, we show that GAS2L2 is abundant at the apical surface of ciliated cells, where it localizes with basal bodies, basal feet, rootlets, and actin filaments. Cultured GAS2L2-deficient nasal epithelial cells from one of the affected individuals showed defects in ciliary orientation and had an asynchronous and hyperkinetic (GAS2L2-deficient = 19.8 Hz versus control = 15.8 Hz) ciliary-beat pattern. These results were recapitulated in Gas2l2-/- mouse tracheal epithelial cell (mTEC) cultures and in X. laevis embryos treated with Gas2l2 morpholinos. In mice, the absence of Gas2l2 caused neonatal death, and the conditional deletion of Gas2l2 impaired mucociliary clearance (MCC) and led to mucus accumulation. These results show that a pathogenic variant in GAS2L2 causes a genetic defect in ciliary orientation and impairs MCC and results in PCD.

Role of Spdef in the Regulation of Muc5b Expression in the Airways of Naive and Mucoobstructed Mice.

Understanding how expression of airway secretory mucins MUC5B and MUC5AC is regulated in health and disease is important to elucidating the pathogenesis of mucoobstructive respiratory diseases. The transcription factor SPDEF (sterile α-motif pointed domain epithelial specific transcription factor) is a key regulator of MUC5AC, but its role in regulating MUC5B in health and in mucoobstructive lung diseases is unknown. Characterization of Spdef-deficient mice upper and lower airways demonstrated region-specific, Spdef-dependent regulation of basal Muc5b expression. Neonatal Spdef-deficient mice exhibited reductions in BAL Muc5ac and Muc5b. Adult Spdef-deficient mice partially phenocopied Muc5b-deficient mice as they exhibited reduced Muc5b in nasopharyngeal and airway epithelia but not in olfactory Bowman glands, 75% incidence of nasopharyngeal hair/mucus plugs, and mild bacterial otitis media, without defective mucociliary clearance in the nasopharynx. In contrast, tracheal mucociliary clearance was reduced in Spdef-deficient mice in the absence of lung disease. To evaluate the role of Spdef in the development and persistence of Muc5b-predominant mucoobstructive lung disease, Spdef-deficient mice were crossed with Scnn1b-transgenic (Scnn1b-Tg) mice, which exhibit airway surface dehydration-induced airway mucus obstruction and inflammation. Spdef-deficient Scnn1b-Tg mice exhibited reduced Muc5ac, but not Muc5b, expression and BAL content. Airway mucus obstruction was not decreased in Spdef-deficient Scnn1b-Tg mice, consistent with Muc5b-dominant Scnn1b disease, but increased airway neutrophilia was observed compared with Spdef-sufficient Scnn1b-Tg mice. Collectively, these results indicate that Spdef regulates baseline Muc5b expression in respiratory epithelia but does not contribute to Muc5b regulation in a mouse model of Muc5b-predominant mucus obstruction caused by airway dehydration.

Quantitative Proteomic Analysis of Human Airway Cilia Identifies Previously Uncharacterized Proteins of High Abundance.

Cilia are essential to many diverse cellular processes. Although many major axonemal components have been identified and studied, how they interact to form a functional axoneme is not completely understood. To further our understanding of the protein composition of human airway cilia, we performed a semiquantitative analysis of ciliary axonemes using label-free LC/MSE, which identified over 400 proteins with high confidence. Tubulins were the most abundant proteins identified, with evidence of 20 different isoforms obtained. Twelve different isoforms of axonemal dynein heavy chain were also identified. Absolute quantification of the nontubulin components demonstrated a greater than 75-fold range of protein abundance (RSPH9;1850 fmol vs CCDC103;24 fmol), adding another level of complexity to axonemal structure. Of the identified proteins, ∼70% are known axonemal proteins. In addition, many previously uncharacterized proteins were identified. Unexpectedly, several of these, including ERICH3, C1orf87, and CCDC181, were present at high relative abundance in the cilia. RT-PCR analysis and immunoblotting confirmed cilia-specific expression for eight uncharacterized proteins, and fluorescence microscopy demonstrated unique axonemal localizations. These studies have provided the first quantitative analysis of the ciliary proteome and have identified and characterized several previously unknown proteins as major constituents of human airway cilia.

Cilia and Mucociliary Clearance.

Mucociliary clearance (MCC) is the primary innate defense mechanism of the lung. The functional components are the protective mucous layer, the airway surface liquid layer, and the cilia on the surface of ciliated cells. The cilia are specialized organelles that beat in metachronal waves to propel pathogens and inhaled particles trapped in the mucous layer out of the airways. In health this clearance mechanism is effective, but in patients with primary cilia dyskinesia (PCD) the cilia are abnormal, resulting in deficient MCC and chronic lung disease. This demonstrates the critical importance of the cilia for human health. In this review, we summarize the current knowledge of the components of the MCC apparatus, focusing on the role of cilia in MCC.

A grafted ovarian fragment rescues host fertility after chemotherapy.

STUDY QUESTION: Can host fertility be rescued by grafting of a fragment of a healthy ovary soon after chemotherapy? SUMMARY ANSWER: We found that grafting a green fluorescent protein (GFP)-positive fragment from a healthy isogenic ovary to the left ovary of a chemo-treated host rescued function and fertility of the grafted host ovary, and resulted in the production of host-derived offspring as late as the sixth litter after chemotherapy (CTx) treatment, whereas none of the ungrafted controls produced a second litter. WHAT IS KNOWN ALREADY: In women and girls undergoing chemotherapy, infertility and premature ovarian failure are frequent outcomes. There are accumulating reports of improved endocrine function after autotransplantation of an ovarian fragment, raising the possibility that the transplant is beneficial to the endogenous ovary. STUDY DESIGN, SIZE, DURATION: We first established a CTx treatment regimen that resulted in the permanent loss of fertility in 100% of female mice of the FVB inbred strain. We grafted an isogenic ovary fragment from a healthy female homozygous for a GFP transgene to the left ovary of 100 CTx-treated hosts, and compared fertility to 39 ungrafted controls in 6 months of continuous matings, using GFP to distinguish offspring derived from the graft, and those derived from the host. PARTICIPANTS/MATERIALS, SETTING, METHODS: Immunofluoresece and western blot analysis of 39 treated ovaries during and 15 days after CTx treatment revealed elevated apoptosis, rapid loss of granulosa cells and an increased recruitment of growing follicles. Using immunofluorescence and confocal imaging, we tracked the outcome of the grafted tissue over 4 months and its effect on the adjacent and contralateral ovary of the host. MAIN RESULTS AND THE ROLE OF CHANCE: Fifty-three percent of grafted females produced a second litter whereas none of the ungrafted females produced a second litter. The likelihood that this could occur by chance is very low (P < 0.0001). LIMITATIONS, REASONS FOR CAUTION: These results are shown only in mice, and whether or how they might apply to chemotherapy patients subjected to different CTx regimens is not yet clear. WIDER IMPLICATIONS OF THE FINDINGS: Our experiments prove that rescue of a chemo-treated ovary is possible, and establish a system to investigate the mechanism of rescue and to identify the factors responsible with the long-term goal of developing therapies for preservation of ovarian endocrine function and fertility in women undergoing chemotherapy. LARGE SCALE DATA: No large datasets were produced. STUDY FUNDING/COMPETING INTERESTS: Duke University Medical Center Chancellor's Discovery Grant to BC; ESJ was supported by an NRSA 5F31CA165545; SK was supported by NIH RO1 GM08033; RWT was supported by the Duke University School of Medicine Ovarian Cancer Research Fellowship; XBM was supported by CONICYT. The authors have no conflicts of interest to declare.

Left-Biased Spermatogenic Failure in 129/SvJ Dnd1Ter/+ Mice Correlates with Differences in Vascular Architecture, Oxygen Availability, and Metabolites.

Homozygosity for the Ter mutation in the RNA-binding protein Dead end 1 (Dnd1(Ter/Ter)) sensitizes germ cells to degeneration in all mouse strains. In 129/SvJ mice, approximately 10% of Dnd1(Ter/+) heterozygotes develop spermatogenic failure, and 95% of unilateral cases occur in the left testis. The first differences between right and left testes were detected at Postnatal Day 15 when many more spermatogonial stem cells (SSCs) were undergoing apoptosis in the left testis compared to the right. As we detected no significant left/right differences in the molecular pathway associated with body axis asymmetry or in the expression of signals known to promote proliferation, differentiation, and survival of germ cells, we investigated whether physiological differences might account for asymmetry of the degeneration phenotype. We show that left/right differences in vascular architecture are associated with a decrease in hemoglobin saturation and increased levels of HIF-1alpha in the left testis compared to the right. In Dnd1 heterozygotes, lower oxygen availability was associated with metabolic differences, including lower levels of ATP and NADH in the left testis. These experiments suggest a dependence on oxygen availability and metabolic substrates for SSC survival and suggest that Dnd1(Ter/+) SSCs may act as efficient sensors to detect subtle environmental changes that alter SSC fate.

Testicular teratomas: an intersection of pluripotency, differentiation and cancer biology.

Teratomas represent a critical interface between stem cells, differentiation and tumorigenesis. These tumors are composed of cell types representing all three germ layers reflecting the pluripotent nature of their cell of origin. The study of these curious tumors became possible when Leroy Stevens identified the 129 mouse strain as a model of spontaneous testicular teratoma and later isolated a substrain carrying the Ter mutation, a potent modifier of tumor incidence. Early studies with 129 mice lead to the discovery of embryonal carcinoma (EC) cells which played a foundational role in the embryonic stem (ES) cell field and the study of pluripotency. The cells of origin of testicular teratomas are germ cells. During early development, primordial germ cells diverge from somatic differentiation and establish their pluripotent nature, maintaining or re-expressing core pluripotency genes; Oct4, Sox2 and Nanog. It is believed that a misregulation of male germ cell pluripotency plays a critical role in teratoma development. Several mouse models of teratoma development have now been identified, including a chromosome substitution strain, 129-Chr19(MOLF), conditional Dmrt1 and Pten alleles and the Ter mutation in the Dnd1 gene. However, it is still unknown what role somatic cells and/or physiology play in the sensitivity to teratoma development. These unusual tumors may hold the key to understanding how pluripotency is regulated in vivo.

Apoptosis, necrosis and autophagy are influenced by metabolic energy sources in cultured rat spermatocytes.

Apoptosis, necrosis and autophagy are mechanistically related processes that control tissue homeostasis and cell survival. In the testis, germ cell death is important for controlling sperm output, but it is unknown whether or not germ cells can switch from apoptosis to necrosis, as has been reported in other tissues. Furthermore, autophagy has not been reported in spermatogenesis. Spermatocytes (meiotic cells) and spermatids (haploid cells) use lactate rather than glucose as their primary substrate for producing ATP. The metabolism of glucose, but not lactate, reduces ATP levels and increases intracellular [H(+)] and [Ca(2+)], both of which are associated with apoptosis and/or necrosis in somatic cells. In this work, we evaluated whether different energy sources, such as lactate or glucose, can influence spermatocyte death type and/or survival in primary cultures. Spermatocytes cultured for 12 h without an energy source died by necrosis, while spermatocytes cultured with 5 mM glucose showed a significant increase in apoptosis, as evidenced by caspase activity, TUNEL assay and phosphatidylserine exposure. Apoptosis was not observed in spermatocytes cultured with 5 mM lactate or deoxyglucose. Autophagy markers, such as LC3-II and autophagosomes, were detected after 12 h of culture, regardless the culture conditions. These results suggest that the availability of glucose and/or lactate affect the type of death or the survival of primary spermatocytes, where glucose can induce apoptosis, while lactate is a protective factor.

TACE/ADAM17 is involved in germ cell apoptosis during rat spermatogenesis.

The pathways leading to male germ cell apoptosis in vivo are poorly understood, but are highly relevant for the comprehension of sperm production regulation by the testis. In this work, we show the evidence of a mechanism where germ cell apoptosis is induced through the inactivation and shedding of the extracellular domain of KIT (c-kit) by the protease TACE/a disintegrin and metalloprotease 17 (ADAM17) during the first wave of spermatogenesis in the rat. We show that germ cells undergoing apoptosis lacked the extracellular domain of the KIT receptor. TACE/ADAM17, a membrane-bound metalloprotease, was highly expressed in germ cells undergoing apoptosis as well. On the contrary, cell surface presence of ADAM10, a closely related metalloprotease isoform, was not associated with apoptotic germ cells. Pharmacological inhibition of TACE/ADAM17, but not ADAM10, significantly prevented germ cell apoptosis in the male pubertal rat. Induction of TACE/ADAM17 by the phorbol-ester phorbol 12-myristate 13-acetate (PMA) induced germ cell apoptosis, which was prevented when an inhibitor of TACE/ADAM17 was present in the assay. Ex-vivo rat testis culture showed that PMA induced the cleavage of the KIT extracellular domain. Isolation of apoptotic germ cells showed that even though protein levels of TACE/ADAM17 were higher in apoptotic germ cells than in nonapoptotic cells, the contrary was observed for ADAM10. These results suggest that TACE/ADAM17 is one of the elements triggering physiological germ cell apoptosis during the first wave of spermatogenesis.